Internal Transcribed Spacer Phylogeny Research Articles

Northward migration past the nearctic biogeographical divide; neotropical Gyrodactylus spp. infecting Astyanax have crossed the Trans-Mexican Volcanic Belt

The neotropical fish genus Astyanax (Characidae) and its associated helminths migrated northward from South America following the Great American Biotic Interchange (GABI): ca. 150 Astyanax spp. are found throughout South and Central America, up to the Mexico-USA border. Most characids are distributed south of the Trans-Mexican Volcanic Belt (TMVB), which bisects the country and represents a major transition zone between the neotropical and nearctic realms. Here, we characterize parasites of the monogenean genus Gyrodactylus infecting Astyanax spp. in Mexico: Astyanax aeneus south of the TMBV, Astyanax mexicanus north of it. Based on morphological, phylogenetic (internal transcribed spacer (ITS) and cytochrome oxidase subunit II (cox 2)) and statistical analyses of morphometric data, we confirmed the validity of Gyrodactylus pakan and Gyrodactylus teken, and erected two new species, Gyrodactylus aphaa n. sp. and Gyrodactylus ricardoi n. sp. These four gyrodactylids are part of a complex of morphologically cryptic species, which are phylogenetically closely related to each other, and sister species to Gyrodactylus carolinae and Gyrodactylus heteracanthus, parasites of characins in Brazil. Four gyrodactylid lineages (G. pakan, G. ricardoi n. sp., G. teken, Gyrodactylus sp. A) are distributed north of the TMVB; G. pakan is also widely distributed south of the TMVB, together with G. aphaa n. sp. Based on the ITS phylogeny, Brazilian parasites form a sister clade to all Mexican gyrodactylids, whose derived clades are distributed in progressively more northerly latitudes in Mexico – the three most-derived species north of the TMVB. This would suggest that gyrodactylid species diverged gradually, presumably as their characid fish hosts colonized and adapted to new environments north of the TMVB.

Two novel members of Onygenales, Keratinophyton kautmanovae and K. keniense spp. nov. from soil

Two new Keratinophyton species, K. kautmanovae sp. nov. and K. keniense sp. nov., isolated from soil samples originating from two different geographical and environmental locations (Africa and Europe) are described and illustrated. Phylogenetically informative sequences obtained from the internal transcribed spacer (ITS) region and the nuclear large subunit (LSU) rDNA, as well as their unique phenotype, fully support novelty of these two fungi for this genus. Based on ITS and LSU combined phylogeny, both taxa are resolved in a cluster with eight accepted species, including K. alvearium, K. chongqingense, K. hubeiense, K. durum, K. lemmensii, K. siglerae, K. submersum, and K. sichuanense. The new taxon, K. kautmanovae, is characterized by clavate, smooth to coarsely verrucose conidia, absence of arthroconidia, slow growth at 25 °C, and no growth at 30 °C, while K. keniense is morphologically unique with a high diversity of conidial shapes (clavate, filiform, globose, cymbiform and rhomboid). Both species are described based on their asexual, a chrysosporium-like morph. While the majority of hitherto described Keratinophyton taxa came from Europe, India and China, the new species K. keniense represents the first reported taxonomic novelty for this genus from Africa.

Identification and Characterization of Pythium deliense Causing Sugar Beet (Beta vulgaris) Root Rot in Grand Forks, North Dakota

Damping-off disease caused by multiple species of soilborne pathogens causes severe economic losses to sugar beet production. Thus, identifying the causal agent is important to formulating effective disease management. During a July 2021 disease survey in a sugar beet field at Grand Forks, North Dakota, we observed that approximately 15% of plants located in low areas of a commercial field showed wilting leaves that were pale green and slightly blighted in color, and the roots were dark brown to black, along with irregular necrotic lesions. The isolated pathogen had the characteristic morphology of Pythium deliense. The sequences of the internal transcribed spacer (ITS) region and the cytochrome c oxidase subunit 1 mitochondrial ( cox1) genes were 100% homologous to corresponding sequences in the NCBI database, which confirmed its P. deliense identity. Moreover, the ITS phylogeny confirmed its identity as P. deliense and its close relationship with P. aphanidermatum. A disease screening trial with four sugar beet varieties was done. Growers and breeders could benefit from accurate information about the sugar beet pathogen in a given field to formulate effective disease management practices.

Validation of Fuscoporia (Hymenochaetales, Basidiomycota) ITS sequences and five new species based on multi-marker phylogenetic and morphological analyses

Although there is a continuous increase in available molecular data, not all sequence identities in public databases are always properly verified and managed. Here, the sequences available in GenBank for Fuscoporia (Hymenochaetales) were validated. Many morphological characters of Fuscoporia overlap among the species, emphasizing the role of molecular identification for accuracy. The identities of 658 Fuscoporia GenBank internal transcribed spacer (ITS) sequences were assessed using ITS phylogeny, revealing 109 (16.6%) misidentified and 196 (29.8%) unspecified sequences. They were validated and re-identified based on the research articles they were published in and, if unpublished, based on sequences from the type, type locality-derived sequences, or otherwise reliable sequences. To enhance the resolution of species delimitation, a phylogenetic assessment of a multi-marker dataset (ITS + nrLSU + rpb2 + tef1) was conducted. The multi-marker phylogeny resolved five of the twelve species complexes found in the ITS phylogeny and uncovered five new Fuscoporia species: F. dolichoseta, F. gilvoides, F. koreana, F. reticulata, and F. semicephala. The validated ITS sequences in this study may prevent further accumulation of misidentified sequences in public databases and contribute to a more accurate taxonomic evaluation of Fuscoporia species.

Potential toxicity of wild Ipomoea ingested by schoolchildren in remote Northeastern Thailand

Background: Natural plant toxins can cause food poisoning upon intentional or unintentional consumption of wild plants. Some toxic wild plants can be mistaken for edible species because of their morphological resemblance. This study examined a poisoning case report of schoolchildren who consumed a steamed tuberous root of wild Ipomoea, misidentified as I. mauritiana, and experienced gastrointestinal toxicity. Objectives: This study aimed to identify the tuberous root of wild Ipomoea using the internal transcribed spacer (ITS) region as a DNA barcode and characterize compounds obtained using gas chromatography-mass spectrometry (GC-MS). Materials and methods: DNA was extracted from fresh and cooked samples of the storage root. PCR amplification and DNA sequencing of the entire ITS region were performed. FastTree and maximum likelihood analyses were used to obtain phylogenetic trees of the Ipomoea species. Root extracts were prepared for GC-MS analysis, and potentially harmful phytochemicals responsible for poisonous plant exposure were predicted based on a well-established plant toxin database. Results: ITS phylogeny showed a close relationship between wild toxic Ipomoea and edible I. mauritiana. The chemometric profile obtained from GC-MS analysis of the root extracts revealed the presence of 31 phytochemicals. Among them, two putatively toxic compounds identified were β-amyrin and coumarin. Conclusion: Misidentification of the wild poisonous plant reported herein resulted in toxic plant ingestion. Although most poisonous plant exposures are not life threatening, measures should be taken to ensure the safety of the general public.

Two new species of Phaeohelotium (Leotiomycetes: Helotiaceae) from Chile and their putative ectomycorrhizal status.

Species of the genus Phaeohelotium (Leotiomycetes: Helotiaceae) are cup fungi that grow on decaying wood, leaves, litter, and directly on soil. Northern Hemisphere species are primarily found on litter and wood, whereas in the Southern Hemisphere the genus includes a mix of saprotrophs as well as taxa that grow on soil in association with ectomycorrhizal trees. The diversity of this genus has not been fully explored in southern South America. Here we describe two species from Chile, Phaeohelotium maiusaurantium sp. nov. and Ph. pallidum sp. nov., found on soil in Patagonian Nothofagaceae-dominated forests. We present macro- and micromorphological descriptions, illustrations, and molecular phylogenetic analyses. The two new species are placed in Phaeohelotium with high support in our 15-locus phylogeny as well as phylogenetic reconstructions based on the internal transcribed spacer (ITS) region of the nuclear ribosomal RNA gene. Our ITS phylogeny places both Ph. maiusaurantium and Ph. pallidum in a well-supported subclade that includes ectomycorrhizal root tip samples from Australasia. Similar species can be separated from these new taxa based on morphological characteristics, biogeography, substrate, and sequence data. In addition, two unnamed species from Chilean Nothofagaceae forests (Phaeohelotium sp. 1 and Phaeohelotium sp. 2) are documented from scant collections and sequence data and await description until more material becomes available. Citation: Grupe II AC, Smith ME, Weier A, Healy R, Caiafa MV, Pfister DH, Haelewaters D, Quandt CA (2022). Two new species of Phaeohelotium (Leotiomycetes: Helotiaceae) from Chile and their putative ectomycorrhizal status. Fungal Systematics and Evolution 10: 231-249. doi: 10.3114/fuse.2022.10.10.

P393 Clinical spectrum and molecular characterization of Rhytidhysteron spp. (now Gloniopsis) from India

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PMObjective Rhytidhysteron spp. are emerging dematiaceous fungi implicated in subcutaneous infections in immunocompromised as well as immunocompetent individuals. Diagnosis of Rhytidhysteron spp is delayed owing to non-sporulation and misidentification. The present study was conducted to characterize Rhytidhysteron spp. isolated from India by molecular techniques.MethodWe retrieved a total of eight isolates of Rhytidhysteron spp from our National Culture Collection for Pathogenic Fungi (NCCPF) that were received from various centers across India. Colony morphology on Sabouraud's dextrose agar (SDA) and potato dextrose agar (PDA) was recorded. Molecular identification was performed by sequencing internal transcribed spacer (ITS) and 28 s region of rDNA. Phylogenetic tree was constructed using the Neighbor-Joining method along with sequences of standard strains retrieved from the NCBI database.Results Rhytidhysteron spp was confirmed by sequencing from cutaneous specimens of eight cases. Six patients were immunocompetent while two were post-renal transplant recipients. Phenotypic features on SDA and PDA revealed growth of greyish-black mycelia on the obverse with black pigmentation on reverse after 7 days of incubation. On microscopic examination of lactophenol cotton blue mount, only dematiaceous septate hyphae without any spores were noted. On phylogenetic analysis of 28 s region, four of our isolates were closely clustered with Gloniopsis calami (Fig.1b), whereas two isolates with G. pneumonia and one each with G. percutanea, and Gloniopsis spp. (Fig.1b). On ITS phylogeny, the same four isolates closely clustered with G. calami (Fig.1a) as in 28 s region, one each isolate clustered with G. pneumonia, and G. percutanea while two isolates formed a separate clade with Rhytidhysteron rufulum (Fig.1a).ConclusionThis fungus is often difficult to identify due to lack of sporulation making morphological identification challenging. Therefore, molecular sequencing is a must for its identification. However, identification using the sequence of ITS and 28 s rDNA does not clearly correlate. Hence to confirm the identity additional genes needs to be sequenced and analyzed.

Molecular detection of a novel perkinsid associated with the deep-sea clam Phreagena okutanii.

Based on environmental DNA surveys, it is widely held that phylogenetically diverse protists exist in chemosynthetic ecosystems. However, knowledge regarding the protists associated with the endemic animals inhabiting these environments is still very limited. In the present study, utilizing polymerase chain reaction (PCR) techniques, we detected fragments of the small subunit ribosomal RNA (SSU rRNA) gene and the internal transcribed spacer (ITS) region of the ribosomal RNA genes from a particular protist in the gills of the vesicomyid clam Phreagena okutanii (formerly described as Calyptogena okutanii), a representative animal in chemosynthetic ecosystems. Based on the phylogeny of the SSU rRNA gene, the organism in question belongs to the genus Perkinsus, which is exclusively composed of protistan parasites infecting mollusks. Intriguingly, based on the ITS phylogeny, this protist was not related to any known Perkinsus species and was deeply branched within the radiation of this genus, thus represents an undescribed species. In addition, the protist detected by PCR was localized to the intercellular spaces in the gills of the host clam with fluorescence in situ hybridization. Although the ecological significance of this novel deep-sea perkinsid remains unclear, our present findings may provide important insights into the diversity of the genus Perkinsus.

Complete Chloroplast Genome of Cnidium monnieri (Apiaceae) and Comparisons with Other Tribe Selineae Species

Cnidium monnieri is an economically important traditional Chinese medicinal plant. In this study, the complete chloroplast (cp) genome of C. monnieri was determined using the Illumina paired-end sequencing, the GetOrganelle de novo assembly strategy, as well as the GeSeq annotation method. Our results showed that the cp genome was 147,371 bp in length with 37.4% GC content and included a large single-copy region (94,361 bp) and a small single-copy region (17,552 bp) separated by a pair of inverted repeat regions (17,729 bp). A total of 129 genes were contained in the cp genome, including 85 protein-coding genes, 36 tRNA genes, and eight rRNA genes. We also investigated codon usage, RNA editing, repeat sequences, simple sequence repeats (SSRs), IR boundaries, and pairwise Ka/Ks ratios. Four hypervariable regions (trnD-trnY-trnE-trnT, ycf2, ndhF-rpl32-trnL, and ycf1) were identified as candidate molecular markers for species authentication. The phylogenetic analyses supported non-monophyly of Cnidium and C. monnieri located in tribe Selineae based on the cp genome sequences and internal transcribed spacer (ITS) sequences. The incongruence of the phylogenetic position of C. monnieri between ITS and cpDNA phylogenies suggested that C. monnieri might have experienced complex evolutions with hybrid and incomplete lineage sorting. All in all, the results presented herein will provide plentiful chloroplast genomic resources for studies of the taxonomy, phylogeny, and species authentication of C. monnieri. Our study is also conducive to elucidating the phylogenetic relationships and taxonomic position of Cnidium.

Hybrid origin of the invasive Spartina anglica inferred from chloroplast and nuclear ITS phylogenies

Spartina anglica originated from interspecific hybridization between S. alterniflora and S. maritima which produced a sterile hybrid S. × townsendii. Subsequent whole genome duplication of the hybrid, gave rise to a vigorous allopolyploid S. anglica. In earlier studies, both parental nuclear sequences were detected in S. anglica. However, to date, parental copies of nuclear ribosomal DNA (nrDNA) have not been detected in a phylogenetic context. In the present study, we used the nuclear internal transcribed spacer (ITS) region in conjunction with chloroplast DNA (trnL-trnF, matK, and rbcL) to infer the hybrid origin of S. anglica invading Korean coasts. Our results revealed that the chloroplast DNA sequences of S. anglica were similar to those of S. alterniflora, which confirms its maternal origin. However, in the nuclear rDNA ITS region, we detected two highly divergent groups of ITS sequences that were divided between the two parent lineages. The maternal copies were indistinguishable from S. alterniflora, while the paternal repeats were highly differentiated from S. maritima and exhibited characteristics of ITS pseudogenes. This indicates that S. anglica has undergone rapid changes at the rDNA loci after the allopolyploid formation. Our study demonstrated the two divergent copies of ITS sequences, thus resolving the phylogenetic hybrid origin of S. anglica. This study offers further insights into the evolutionary history of S. anglica.

Phylogenetic origin of two Japanese Torreya taxa found in two regions with strongly contrasting snow depth.

The Japanese archipelago exhibits a notable difference in snow depth in winter, deep snow on the Sea of Japan side and low snow cover on the Pacific Ocean side. This contrasting pattern has shaped the distribution of infraspecific taxon pairs in a range of woody plants, with taxa found on the Sea of Japan side typically exhibiting a stunted shrub form with multiple decumbent stems. The phylogenetic origin of these taxon pairs is unknown, i.e., whether the two taxa diverged from the same species or if they have different origins. This study aimed to reveal the phylogenetic origin of two varieties of Torreya nucifera (Taxaceae); var. nucifera is a tree found on the Pacific Ocean side, whereas var. radicans is a shrub found on the Sea of Japan side. We examined the phylogenetic relationships of the two varieties and worldwide Torreya taxa using whole chloroplast genomes, chloroplast DNA fragments, and the nuclear ribosomal internal transcribed spacer (ITS). The whole chloroplast genome phylogeny indicated that T. nucifera var. radicans was a sister taxon to Chinese T. grandis, rather than to var. nucifera. In contrast, the nuclear ITS phylogeny indicated that while several haplotypes of T. nucifera var. radicans were closely related to T. grandis, most haplotypes of T. nucifera var. radicans formed a single clade with those of var. nucifera. This implies that the homogenization of the ITS has occurred between the two taxa, while taxon-specific chloroplast DNA haplotypes were retained. These discordant phylogenies suggested that the two taxa have different phylogenetic origins, but have an intricate evolutionary history, involving inter-taxa hybridization and gene flow, possibly when their distributions were confined to sympatric refugia. Given the genetic evidence and distinct difference in growth form, we propose that T. nucifera var. radicans should be taxonomically treated as a distinct species, T. fruticosa.

Identification and Characterization of Ceratocystis fimbriata Causing Lethal Wilt on the Lansium Tree in Indonesia

Bark canker, wood discoloration, and wilting of the duku tree (Lansium domesticum) along the watershed of Komering River, South Sumatra Province, Indonesia first appeared in 2013. The incidence of tree mortality was 100% within 3 years in badly infected orchards. A Ceratocystis species was consistently isolated from the diseased tissue and identified by morphological and sequence analyses of the internal transcribed spacer (ITS) and β-tubulin regions. Pathogenicity tests were conducted and Koch’s postulates were confirmed. The fungus was also pathogenic on Acacia mangium, but was less pathogenic on mango. Partial flooding was unfavourable for disease development. Two described isolates (WRC and WBC) had minor variation in morphology and DNA sequences, but the former exhibited a more pathogenic on both duku and acacia. The ITS phylogenies grouped the most pathogenic isolate (WRC) causing wilting of the duku tree within the aggressive and widely distributed ITS5 haplotype of C. fimbriata.

Fins infestation induced by Myxobolus xiantaoensis in yellow catfish Tachysurus fulvidraco Richardson, 1846: Some pathophysiological and molecular insights

The myxozoan parasite Myxobolus xiantaoensis is a fin pathogen of commercially important yellow catfish Tachysurus fulvidraco Richardson, 1846, in the freshwater ponds of China. In the present work, four geographical isolates of M. xiantaoensis were sampled from the fins of yellow catfish. It was found that the spores of four isolates exhibited few markable differences in morphometrics. The small subunit ribosomal DNA (SSU rDNA) sequences of four isolates were conspecific to the SSU rDNA sequence of M. xiantaoensis. No genetic level variation was observed, even in the characteristically more variable internal transcribed spacer (ITS) region. This absence of variability suggests high gene flow as a result of panmixia in the parasitic populations. ITS phylogeny placed four isolates of M. xiantaoensis in a clade together with myxozoans species infecting Siluriformes. The M. xiantaoensis infection inflicted severe hemorrhages on epidermis of ray-fins, which grew into inflammatory epithelial hyperplasia and lytic cartilage signs. The histochemical analysis of infected fins biopsies is characterized by damage of collagen components of cartilage, resulting in weakness, breaks, and missing fin rays. These tissue sections also had a remarkable inflammatory response around the fin cartilage, with the absence of mature spores and chondrocytes. These results indicate that the fin cartilage damage appeared before the development of tissue inflammation and the parasitic infestation of the fins. The present four geographical isolates of M. xiantaoensis were identified by a holistic approach of species characterization based on biological, morphological, and molecular evidence. These four isolates showed some morphological and genetic variations but within the intraspecific range.

Molecular prospecting for cryptic species of the Hypholoma fasciculare complex: toward the effective and practical delimitation of cryptic macrofungal species

Many macrofungal cryptic species remain unidentified. A possible solution is to increase the number of loci analyzed and use rigorous statistics for macrofungal species delimitation. To validate this assumption, cryptic species of the Hypholoma fasciculare complex, a group of common wood-decomposing fungi, were attempted to be delineated. Massively parallel sequencing of mitochondrial ribosomal RNA (mt_rRNA), nuclear ribosomal internal transcribed spacer (ITS) region, and 24 single-copy genes were performed for 96 specimens collected in Japan. Then, the species boundaries were inferred using comparative gene genealogies (mt_rRNA vs. ITS), Bayesian Poisson tree process (bPTP) model for the phylogeny of concatenated nuclear sequences, and analysis of molecular variance (AMOVA) for single nucleotide polymorphisms. In both the mt_rRNA and ITS phylogenies, the H. fasciculare complex was not divided into well-supported clades. Nevertheless, based on the bPTP, two mitochondrial haplotypes were inferred to represent distinct species (H. fasciculare and H. subviride). The results of AMOVA also indicated that the differentiation of nuclear loci can be explained mostly by differences between haplotype. These results suggest that it is necessary to increase the number of target loci to 20 or more and use both phylogeny-based and population genetics-based statistics for the accurate delimitation of macrofungal species.

Species composition, diversity, and distribution of the genus Ulva along the coast of Jeju Island, Korea based on molecular phylogenetic analysis.

Species diversity in the genus Ulva remains understudied worldwide. Using molecular analyses we investigated the species composition, diversity, distribution, and relative frequencies of the genus Ulva along the entire coast of Jeju Island, off the southern tip of Korea. Species identification was performed for 215 samples collected from 23 sites, based on comprehensive phylogenetic and model-based species delimitation analyses using the sequences of two molecular markers, chloroplast elongation factor Tu (tufA) and nuclear rDNA internal transcribed spacer (ITS). We identified 193 specimens as nine Ulva species, 14 specimens as Blidingia spp., and eight samples undetermined, based on the combined analysis of tufA and ITS phylogenies. Two model-based approaches generally supported nine groups of Ulva species. Previously documented species complex, such as U. ohnoi−U. spinulosa and U. procera−U. linza showed discordant relationships between the two phylogenies. The occurrence of U. torta on Jeju Island was first observed, despite its existence on the mainland previously reported. Ulva australis [16 of 23 sites; 34.4% (relative frequency)], U. ohnoi (16; 21.9%), and U. procera (11; 14%) were found to be the predominant species. Our study highlights that molecular analysis is critical for species delimitation in the genus Ulva and provides fundamental information for an understanding of green-tide assemblages on the “biological hotspot” coastal ecosystem, Jeju Island in Korea. This study will also help to monitor and manage local green tides at the areas that are currently encountering rapid climate changes.

Morphological variation, ultrastructure, pigment composition and phylogeny of the star-shaped dinoflagellate Asterodinium gracile (Kareniaceae, Dinophyceae)

ABSTRACTMorphological variation, ultrastructure, pigment composition and phylogeny of an oceanic, star-shaped, unarmored dinoflagellate Asterodinium gracile were examined using a culture established from Nagasaki, Japan, in 2017. Our examinations were based on LM, SEM, TEM, high-performance liquid chromatography (HPLC) and molecular phylogeny inferred from nuclear-encoded internal transcribed spacer (ITS) and 28S rDNA and chloroplast-encoded 16S rDNA sequences. Typical cells were stellate, possessing an apical, two lateral, and two antapical extensions. Cells ranged in size from 62.5 to 105.6 µm long and 21.2 to 39.5 µm in cingular width. The length and orientation of cellular extensions were variable; that is, two lateral extensions were usually symmetrically bent or retracted. Ellipsoidal cells without extensions were also observed. The apical structure complex was straight or slightly sigmoid. A spherical nucleus was located at the left side in the hyposome. The numerous discoid, yellowish-brown chloroplasts were variable in size and distributed also in the cell extensions. Among Kareniaceae, the lack of a pyrenoid in Asterodinium is unique. Nuclear-encoded ITS and 28S phylogenies suggested that A. gracile is closely related to Karenia papilionacea in the Kareniaceae, but its affinity to Brachidinium capitatum, another species with extensions, was unclear. A phylogeny of 16S rDNA showed that chloroplasts of A. gracile are related to other kareniaceans derived from a haptophyte. These chloroplasts were of the fucoxanthin type, composed of pigments similar to those in other Kareniaceae; however, the detection of 19′-hexanoyloxyfucoxanthin as a major carotenoid, the low amount of fucoxanthin, and lack of 19′-butanoyloxyfucoxanthin in A. gracile were unusual for the family.

A re-investigation of the bloom-forming unarmored dinoflagellate Karenia longicanalis (syn. Karenia umbella ) from Chinese coastal waters

The dinoflagellate genus Karenia is known for recurrent harmful blooms worldwide. However, species diversity of the genus is generally overlooked owing to the difficulty of identifying small unarmored dinoflagellates. We have established four clonal cultures of Karenia longicanalis isolated from the type locality, Hong Kong harbor (strain HK01) and other three locations along the Chinese coasts (strains YB01, DT01, and NJ01). The morphology of the strain was studied by light and scanning electron microscopy (LM and SEM) and the pigment composition analyzed by high-performance liquid chromatography. We provide the first molecular data of K. longicanalis based on the large subunit (LSU) rRNA gene sequence and internal transcribed spacer (ITS). The four strains showed identical LSU rDNA sequences with a similarity of 99.4% to the holotype of Karenia umbella (strain KUTN05) from Australia. In the ITS phylogeny, the sequence of K. umbella branched between the Chinese strains of K. longicanalis. A careful comparison of the morphology of K. longicanalis and K. umbella reveals the similarity in the diagnostic characters. Differences may appear due to the sample treatment for SEM. We conclude that K. umbella is a junior synonym of K. longicanalis.

Phylogenetic relationships in Seslerieae (Poaceae) including resurrection of Psilathera and Sesleriella, two monotypic genera endemic to the Alps

AbstractSesleria (Poaceae) is a species‐rich, taxonomically difficult genus due to high morpho‐anatomical variability and weak differentiation of many species. It is distributed throughout western Eurasia and North Africa with highest taxonomic diversity on the Balkan Peninsula. Of the more than 40 species only two are diploid, all others are tetra‐ to dodecaploid, and many species comprise several ploidy cytotypes. We sequenced the plastid trnL‐ndhF region and produced amplified fragment length polymorphisms (AFLPs) from a comprehensive, ploidy‐cytotyped sampling including 292 populations, and produced nuclear ribosomal internal transcribed spacer (ITS) sequences for a subsample. Our first aim was to determine the relationships among Sesleria and putatively closely related genera. Further, we explored whether diploid S. sphaerocephala and S. ovata should be treated as separate genera Sesleriella and Psilathera as proposed previously, or rather included in Sesleria, following most recent Floras. Finally, we tested previous hypotheses of infrageneric classifications. In the ITS phylogeny tetra‐ to dodecaploid Sesleria s.str. grouped with diploid S. ovata whereas diploid S. sphaerocephala was more distantly related. In contrast, in the plastid phylogeny Sesleria s.str. grouped with S. sphaerocephala, whereas S. ovata was sister to Echinaria. This suggests an allopolyploid origin of Sesleria s.str. with S. sphaerocephala acting as maternal and S. ovata as paternal species. The lack of monophyly of Sesleria s.l. thus supports the recognition of S. ovata and S. sphaerocephala as separate genera Psilathera and Sesleriella. Their segregation, which is also supported by morphology, increases the number of angiosperm genera endemic to the European Alps from three to five. In congruence with the plastid sequences, AFLPs resolved the traditionally recognized sections (sect. Argenteae, sect. Sesleria) as non‐overlapping clusters and identified S. vaginalis as a probably recently evolved intersectional hybrid. Within the two sections, several informal species groups could be recognized, which show limited congruence with those proposed in the most comprehensive taxonomic treatment of the genus. From an evolutionary point of view it is obvious that ancient as well as recent hybridization coupled with polyploidisation have played a crucial role in diversification within Sesleria.

Phylogenetic study of the endemic species Oxytropis almaatensis (Fabaceae) based on nuclear ribosomal DNA ITS sequences

BackgroundOxytropis almaatensis Bajt. is a rare, narrow endemic species of the Trans-Ili Alatau mountains in Kazakhstan. Up to now, no studies regarding the taxonomy and variation of key morphological traits of O. almaatensis were undertaken. The purpose of this analysis was to evaluate phenotypic variation of O. almaatensis and assess the position of the species within the genus based on nucleotide sequences of the nuclear ribosomal DNA internal transcribed spacer (ITS) region.ResultsTwo populations of O. almaatensis were collected in neighboring gorges of the Trans-Ili Alatau Mountains. The ITS sequences from the samples of two populations of O. almaatensis were identical. The phylogenetic analysis indicated that O. almaatensis is within Oxytropis genetically close to O. glabra as these species formed a separate subclade. The phenotypic variation of populations was assessed using nine morphological traits and compared to descriptions of O. glabra. The range of variation for the traits between two populations was established. A clear morphological difference of O. almaatensis and O. glabra was found in peduncle length to leaf length ratio. This was in O. almaatensis 1.56, while in O. glabra, it was 1.0.ConclusionsThe study provides the first phenotypic description and phylogenetic placement of the rare endemic species O. almaatensis. The morphological traits in two O. almaatensis populations showed a high level of phenotypic variability. Although clearly different from O. glabra, the ITS phylogeny grouped these species in a subclade within the genus.

Contribution of dark septate fungi to the nutrient uptake and growth of rice plants.

The use of dark septate fungi (DSE) to promote plant growth can be beneficial to agriculture, and these organisms are important allies in the search for sustainable agriculture practices. This study investigates the contribution of dark septate fungi to the absorption of nutrients by rice plants and their ensuing growth. Four dark septate fungi isolates that were identified by Internal transcribed spacer phylogeny were inoculated in rice seeds (Cv. Piauí). The resulting root colonization was estimated and the kinetic parameters Vmax and Km were calculated from the nitrate contents of the nutrient solution. The macronutrient levels in the shoots, and the NO3−-N, NH4+-N, free amino-N and soluble sugars in the roots, sheathes and leaves were measured. The rice roots were significantly colonized by all of the fungi, but in particular, isolate A103 increased the fresh and dry biomass of the shoots and the number of tillers per plant, amino-N, and soluble sugars as well as the N, P, K, Mg and S contents in comparison with the control treatment. When inoculated with isolates A103 and A101, the plants presented lower Km values, indicating affinity increases for NO3−-N absorption. Therefore, the A103 Pleosporales fungus presented the highest potential for the promotion of rice plant growth, increasing the tillering and nutrients uptake, especially N (due to an enhanced affinity for N uptake) and P.